溶血卵磷脂对牛视网膜微血管内皮细胞转化生长因子-β_2及其Ⅱ型受体表达的影响

目的:探讨不同浓度的溶血卵磷脂(LPC)对牛视网膜微血管内皮细胞转化生长因子-β2(TGF-β2)及其Ⅱ型受体表达的影响.方法:原代分离培养牛的视网膜微血管内皮细胞,加入不同浓度的LPC,分别培养6、12、24h,RT-PCR检测TGF-β2 mRNA表达水平,半巢式RT-PCR检测TGF-βⅡ型受体mRNA的表达水平.结果:LPC的浓度为40 μmol/L作用12 h后,TGF.B2 mRNA水平明显高于浓度为20 μmol/L和60 μmol/L.LPC浓度改变对TGF-βⅡ型受体mRNA的表达无明显影响.结论:LPC能调节牛视网膜内皮细胞TGF-β2的表达;LPC对TGF-βⅡ型受体的表达无明显影响.     

A phospholipase A2 isolated from Lachesis muta snake venom increases the survival of retinal ganglion cells in vitro

We have previously showed that a phospholipase A₂ isolated from Lachesis muta snake venom and named LM-PLA₂-I displayed particular biological activities, as hemolysis, inhibition on platelet aggregation, edema induction and myotoxicity. In the present work, we evaluated the effect of LM-PLA₂-I on the survival of axotomized rat retinal ganglion cells kept in vitro, as well as its mechanism of action. Our results clearly showed that treatment with LM-PLA₂-I increased the survival of ganglion cells (100% when compared to control cultures) and the treatment of LM-PLA₂-I with p-bromophenacyl bromide abolished this effect. This result indicates that the effect of LM-PLA₂-I on ganglion cell survival is entirely dependent on its enzymatic activity and the generation of lysophosphatidylcholine (LPC) may be a prerequisite to the observed survival. In fact, commercial LPC mimicked the effect of LM-PLA₂-I upon ganglion cell survival. To investigate the mechanism of action of LM-PLA₂-I, cultures were treated with chelerythrine chloride, BAPTA-AM, rottlerin and also with an inhibitor of c-junc kinase (JNKi). Our results showed that rottlerin and JNK inhibitor abolished the LM-PLA₂-I on ganglion cell survival. Taken together, our results showed that LM-PLA₂-I and its enzymatic product, LPC promoted survival of retinal ganglion cells through the protein kinase C pathway and strongly suggest a possible role of the PLA₂ enzyme and LPC in controlling the survival of axotomized neuronal cells.     

Down-regulation of endogenous nitric oxide synthase inhibitors on endothelial SK3 expression

OBJECTIVES: To investigate role of endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine (ADMA) in down-regulation of the expression of endothelial SK3 in atherosclerosis. METHODS: Apolipoprotein E deficient (apo E(-/-)) mice aged 11 approximately 12 weeks were treated with ADMA (5 mg/kg per day, subcutaneous injection) for 4 weeks. Cultured human umbilical venous endothelial cells (HUVECs) were treated with different concentrations of lysophosphatidylcholine (LPC) or ADMA for 48 h. Plasma levels of ADMA were determined by high performance liquid chromatogram (HPLC); protein and mRNA levels of SK3 in the aortas of mice and cultured cells were detected by immunofluorescence, western blot and RT-PCR, respectively. RESULTS: Concomitantly with the elevated plasma levels of ADMA, the expressions of both SK3 protein and mRNA in aortas of apo E(-/-) mice were significantly reduced in comparison to those of the wild-type mice. Moreover, 4-week treatment of ADMA made levels of SK3 expression even lower. In cultured HUVECs, either LPC or ADMA notably decreased the expressions of both SK3 protein and mRNA in a concentration dependent manner. CONCLUSIONS: Endogenous ADMA may be an important factor for down-regulation of the expression of endothelial SK3 in atherosclerotic animals.     

蛋白激酶C激动剂和抑制剂对溶血磷脂酰胆碱引起脑微血管平滑肌细胞增殖的影响

目的：探讨蛋白激酶C（PKC）激动剂与抑制剂对溶血磷脂酰胆碱（LPC）引起脑微血管平滑肌细胞增殖的影响。方法：在建立了LPC引起牛脑微血管平滑肌细胞（BCMSMC）增殖模型基础上，以结晶紫染色法测定PKC激动剂phorbol 12-myristate 13-acetate(PMA)和抑制剂1－（5－isoquninoline sulfonyl)-2-methylpiperatione hydrochloric(H7)对平滑肌细胞增殖的影响。结果：PMA时间依赖性和剂量依赖性地使LPC引起BCMSMC增殖率显增高，H7非常显地降低了LPC引起的BCMSMC增殖率，亦呈时间依赖性和剂量依赖性。结论：LCP对BCMSMC增殖作用与PKC信号转导系统有相关性。     

Extracellulary administered lysophosphatidylcholine causes Ca2+ efflux from freshly isolated adult rat cardiomyocytes

It has previously been reported that ischemia and reperfusion of the heart cause accumulation of lyophosphatidylcholine (LPC) within the myocardium. While it is known that LPC causes the transient increase of intracellular free Ca²⁺ concentration ([Ca²⁺]_i) during contraction of cardiac cells, little is known about the mechanism for decreasing [Ca²⁺]_i in cardiomyocytes during LPC accumulation. Since cumulative elevation in [Ca²⁺]_i leads to irreversible injury to cardiomyocytes, elevated [Ca²⁺]_i must be restored to an unstimulated level to maintain cell functions. In the present study, we therefore examined the effect of LPC on Ca²⁺ efflux from freshly isolated adult rat cardiomyocytes. LPC stimulated the efflux of ⁴⁵Ca²⁺ from the cells in a concentration‐dependent manner (10^–7 M – 10^–5 M). Other lysophospholipids, which are generated from phopholipids of the cell membrane, failed to induce ⁴⁵Ca²⁺ efflux from the cells. Dilazep and K‐7259, which are known to inhibit the increase in [Ca²⁺]_i caused by LPC, likewise reduced ⁴⁵Ca²⁺ efflux caused by LPC addition. Furthermore, the LPC‐stimulated ⁴⁵Ca²⁺ efflux was not affected by removal of extracellular Ca²⁺, but was dependent on the presence of extracellular Na⁺. On the other hand, inhibitors of Na⁺/Ca²⁺ exchange, amiloride and 5‐(N,N‐dimethyl)‐amiloride, inhibited LPC induced ⁴⁵Ca²⁺ efflux. These results suggest that LPC stimulates extracellular Na⁺‐dependent ⁴⁵Ca²⁺ efflux from freshly isolated adult rat cardiomyocytes, probably through Na⁺/Ca²⁺ exchange on the plasma membrane of cells. Received: 3 March 1997, Returned for revision: 7 April 1997, Revision received: 1 August 1997, Accepted: 5 September 1997     

Impact of serial measurements of lysophosphatidylcholine on 28-day mortality prediction in patients admitted to the intensive care unit with severe sepsis or septic shock

Purpose

The purpose of this study is to investigate the effect of serial lysophosphatidylcholine (LPC) measurement on 28-day mortality prediction in patients with severe sepsis or septic shock admitted to the medical intensive care unit (ICU).

Methods

This is a prospective observational study of 74 ICU patients in a tertiary hospital. Serum LPC, white blood cell, C-reactive protein, and procalcitonin (PCT) levels were measured at baseline (day 1 of enrollment) and day 7. The LPC concentrations were compared with inflammatory markers using their absolute levels and relative changes.

Results

The LPC concentration on day 7 was significantly lower in nonsurvivors than in survivors (68.45 ± 42.36 μmol/L and 99.76 ± 73.65 μmol/L; P = .04). A decreased LPC concentration on day 7 to its baseline as well as a sustained high concentration of PCT on day 7 at more than 50% of its baseline value was useful for predicting the 28-day mortality. Prognostic utility was substantially improved when combined LPC and PCT criteria were applied to 28-day mortality outcome predictions. Furthermore, LPC concentrations increased over time in patients with appropriate antibiotics but not in those with inappropriate antibiotics.

Conclusions

Serial measurements of LPC help in the prediction of 28-day mortality in ICU patients with severe sepsis or septic shock.     

Bioactive Lysophosphatidylcholine 16:0 and 18:0 Are Elevated in Lungs of Asthmatic Subjects

Purpose

Asthma is a chronic inflammatory disease of the airways, and is associated with upregulation of phospholipase A₂ (PLA₂), the enzyme that hydrolyzes phosphatidylcholine, producing lysophosphatidylcholine (LPC) and free fatty acids. LPC is a lipid mediator with known pro-inflammatory and pro-atherogenic properties, and is believed to be a critical factor in cardiovascular diseases. We postulate that asthmatic subjects have an elevated content of LPC in the lung lining fluids.

Methods

Eight non-asthmatic controls and seven asthmatic subjects were recruited for broncho-alveolar lavage fluids (BALF) collection for analysis of LPC by high performance liquid chromatography-tandem mass spectrometry.

Results

LPC16:0 and LPC18:0 were significantly elevated in the BALF of asthmatics with impaired lung function characteristic of moderate asthma, but not mild asthma. The increased LPC content in BALF was accompanied by increased PLA₂ activity. Furthermore, qRT-PCR analysis of the BALF cell fraction indicated increased secretory PLA₂-X (sPLA₂-X).

Conclusions

The increased LPC content in the lung lining fluids is a potential critical lipid mediator in the initiation and/or progression of airway epithelial injury in asthma.